Comprehensive investigations of key mitochondrial metabolic changes in senescent human fibroblasts

Abstract

There is a paucity of detailed data related to the effect of senescence on the mitochondrial antioxidant capacity and redox state of senescent human cells. Activities of TCA cycle enzymes, respiratory chain complexes, hydrogen peroxide (H 2 O 2 ), superoxide anions (SA), lipid peroxides (LPO), protein carbonyl content (PCC), thioredoxin reductase 2 (TrxR2), superoxide dismutase 2 (SOD2), glutathione peroxidase 1 (GPx1), glutathione reductase (GR), reduced glutathione (GSH), and oxidized glutathione (GSSG), along with levels of nicotinamide cofactors and ATP content were measured in young and senescent human foreskin fibroblasts. Primary and senescent cultures were biochemically identified by monitoring the augmented cellular activities of key glycolytic enzymes including phosphofructokinase, lactate dehydrogenase, and glycogen phosphorylase, and accumulation of H 2 O 2 , SA, LPO, PCC, and GSSG. Citrate synthase, aconitase, α-ketoglutarate dehydrogenase, succinate dehydrogenase, malate dehydrogenase, isocitrate dehydrogenase, and complex I-III, II-III, and IV activities were significantly diminished in P25 and P35 cells compared to P5 cells. This was accompanied by significant accumulation of mitochondrial H 2 O 2 , SA, LPO, and PCC, along with increased transcriptional and enzymatic activities of TrxR2, SOD2, GPx1, and GR. Notably, the GSH/GSSG ratio was significantly reduced whereas NAD + /NADH and NADP + /NADPH ratios were significantly elevated. Metabolic exhaustion was also evident in senescent cells underscored by the severely diminished ATP/ADP ratio. Profound oxidative stress may contribute, at least in part, to senescence pointing at a potential protective role of antioxidants in aging-associated disease.

The Tricarboxylic Acid Cycle as a Central Regulator of the Rate of Aging: Implications for Metabolic Interventions

“…ATP levels decline linearly with age in animal models and in human calf and heart muscle. [ 4 ] Bioenergetic failure is seen in certain senescent cells, [ 5 ] certain animal models of aging, [ 6 ] and such human age‐related diseases as Alzheimer's, [ 7 ] osteoporosis, [ 8 ] sarcopenia, [ 9 ] heart failure, [ 10 ] glaucoma, [ 11 ] and COPD. [ 12 ] Cellular functioning depends on intact energy generation.…”

Antioxidant activity of the thioredoxin system

“…After Trx is reduced, the disulfide bonds of other proteins are reduced, thus exerting biological activity. In addition, TR can also reduce antioxidants, protein disulfide isomerase (PDI) (Che et al 2020 ; Ogata et al 2022 ), lipid peroxides and H 2 O 2 (Ghneim et al 2022 ). TR plays an antioxidant role through the catalytic regeneration of coenzyme Q10 (Preci et al 2021 ).…”